Acoustic Biosensor for Discrimination of Pathogens according to the Gram Principle

Abstract

The microacoustic methods of biomedical analysis, implemented on piezoelectric crystals and ceramics, are becoming increasingly popular due to the fact of their potential for integration into laboratories-on-a-chip, biochips, and biosensors as functional elements of biosensors. An important stage in diagnostics of infectious diseases is the identification of pathogens. One possible applications of such a sensor is an alternative to the time- and labor-consuming Gram method of discriminating bacteria according to the composition of their cell walls. Thus, bacteria, which in a Gram staining procedure do not decolor after application of the dye solution, are classified as Gram-positive (G(+)). They are surrounded with a thick peptidoglycan layer that is pulpy and dampens acoustic waves. While Gram-negative (G(–)) bacteria, which acquire a red color in a Gram procedure, are covered with a thin and springy layer, demonstrating resonance effects when interacting with acoustic fields. Thus, G(+) and G(–), which are differently colored in Gram procedures, also react differently to an external acoustic field: for G(–) bacteria, this was a sharp decrease in the Q-factor of the “resonator–suspension” system and a shift of the resonance curve to lower frequencies. While for G(+) bacteria, although a certain shift of the resonance curve was also observed, the bandwidth of the resonance curve practically did not change. This effect was studied for L. acidophilus (G(+)) and Escherichia coli (G(–)) bacilli with quarts resonators of 4 MHz, 5 MHz, and 10 MHz. The biosensor was tested using Lactobacillus fermentum, E. coli M-17, Bifidobacterium bifidum, Burkholderia cepacian, and Staphylococcus aureus. At this stage, it has been demonstrated that the method is particularly effective for discriminating bacteria of a similar shape, such as, for example, cocci. The discrimination of the Gram factor for cocci and bacilli was less accurate and needs further studies for selection of precise resonance frequencies.